As methods of analysis for detecting or quantifying an analyte in a sample with the utilization of immune response specificity, a variety of methodologies, such as immunodiffusion, enzyme assay, and aggregation, have been put to practical use. In particular, detection methods by means of immunochromatography assay (detection) techniques (a lateral-flow assay, a tangential-flow assay, JP Patent Publication (Kokoku) No. 7-13640B (1995), JP Patent No. 2,131,938, and JP Patent No. 2,890,384) spread rapidly because of the convenience thereof in recent years.
The principles of such detection methods are briefly described below. Many types of commercial kits for immunochromatography assay techniques each comprise a sheet-like solid-phase support. On the sheet-like solid-phase support, the following members are sequentially immobilized from an end thereof in the lengthwise direction: 1. a specimen-supply site; 2. a labeled reagent site that holds on a membrane a labeled reagent (e.g., colloidal gold particles that can be visualized or enzymes) containing a ligand that specifically binds to an analyte (e.g., an antigen) so as to be capable of developing on the membrane; and 3. a capture reagent site to which a capture reagent (e.g., an antibody) has been immobilized for capturing an analyte (e.g., an antigen) or label antibody or a complex of the analyte and the label reagent. In such a kit, a solution is continuously transferred via capillary action.
If a given amount of a sample comprising an analyte-containing specimen suspended therein is supplied to the site to which the analyte has been applied, the sample migrates into the site to which the labeled reagent has been applied while developing on the solid-phase support, and the analyte binds to a labeled reagent to form a complex of the analyte and the label reagent. The complex of the analyte and the label reagent develops on the membrane and migrates into the capture reagent site on the membrane, the complex is captured by the capture reagent immobilized on the solid-phase support, and the complex of the capture reagent, the analyte, and the labeled reagent is formed at the capture reagent site. The presence of the analyte can be determined by detecting the labeled reagent by any technique (in the case of colloidal gold particles that can be visualized, an image of the aggregate thereof is to be detected; in the case of enzymes, color development resulting from the addition of a support is to be detected). FIG. 7 shows a conventional immunochromatography detection apparatus.
Because of its constitution, an immunochromatography detection apparatus involves the supply and the development of a specimen on a solid-phase support. This enables the continuous and automatic performance of the following reactions on the solid-phase support: (A) the reaction wherein an analyte specifically binds to a labeled reagent containing a ligand that specifically binds to the analyte; and (B) the reaction wherein a capture reagent specifically binds to the complex of the analyte and the labeled reagent. Thus, detection methods involving the use of such immunochromatography apparatus are excellent in terms of convenience. In fact, however, many detection processes suffer from the presence of impurities in the analyte-containing specimens. Thus, pretreatments such as separation and extraction of analytes from specimens were required in order to remove such impurities via a variety of chemical or physical treatment prior to the assay. In such a case, pretreatments were often conducted under conditions that were not optimal for the binding between the antigen and the antibody. Accordingly, the specimens were required to be optimized for the binding, following the pretreatments. In order to facilitate specific binding of the analyte to the labeled reagent, a pH level, a salt concentration, and other conditions had to be controlled in the reaction system. Since the optimal reaction conditions such as a pH level or a salt concentration of the reaction (A) were not always consistent with those of the reaction (B), such technique was disadvantageous compared with other detection techniques involving a plurality of separate reactions, such as ELISA, in terms of convenience, sensitivity, and specificity.    Patent Document 1: JP Patent Publication (Kokoku) No. 7-13640B (1995)    Patent Document 2: JP Patent No. 2,131,938    Patent Document 3: JP Patent No. 2,504,923    Patent Document 4: JP Patent No. 2,890,384